pK(a) calculations for class C beta-lactamases: The role of Tyr-150

The Poisson-Boltzmann method was used to compute the pK(a) values of titrat able residues in a set of class C beta-lactamases, In these calculations, t he pK(a) of the phenolic group of residue Tyr150 is the only one to stand o ut with an abnormally low value of 8.3, more than one pK(a) unit lower than the measured reference value for tyrosine in solution. Other important res idues of the catalytic pocket, such as the conserved Lys67, Lys315, His314, and Glu272 (hydrogen-bonded to the ammonium group of Lys315), display norm al protonation states at neutral pH. pK(a) values were also computed in cat alytically impaired beta-lactamase mutants. Comparisons between the relativ e k(cat) values and the Tyr150 pK(a) value in these mutants revealed a stri king correlation. In active enzymes, this pK(a) value is always lower than the solution reference value while it is close to normal in inactive enzyme s. These results thus support the hypothesis that the phenolate form of Tyr 150 is responsible for the activation of the nucleophilic serine, The possi ble roles of Lys67 and Lys315 during catalysis are also discussed, (C) 2000 Wiley-Liss, Inc.